| Size | Price | Stock | Qty |
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| 500mg |
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| 1g |
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| Other Sizes |
| Targets |
5-Aminofluorescein targets the mRNA N6-methyladenosine (m6A) demethylase fat mass and obesity-associated (FTO) protein with an IC50 of 6.55 μM. It inhibits the N-terminal 31 residues truncated FTO demethylase activity using m6A-containing ssRNA as a substrate. The mechanisms behind these activities were elucidated after determining the X-ray crystal structures of FTO/fluorescein and FTO/5-aminofluorescein. It is also used as a fluorescent dye for labeling biomolecules.
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| ln Vitro |
In vitro, 5-Aminofluorescein is used as a fluorescent probe to label proteins and other biomolecules. It exhibits excitation and emission wavelengths of 490 nm and 515 nm, respectively. It is chemically coupled to target molecules such as proteins, nucleic acids, or other biomolecules for detection purposes. It has been shown to selectively inhibit FTO demethylation. The IC50 value of 6550.0 nM (6.55 μM) was determined by an assay using LC-MS analysis over a 1-hour incubation period.
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| ln Vivo |
In vivo applications of 5-Aminofluorescein are primarily as a fluorescent marker for imaging and tracking. It is used as a targeted prodrug, suggesting potential for site-specific drug delivery. It has been conjugated to human serum albumin for fluorescence imaging of glioblastomas. These inhibitors can also be applied to the direct labeling and enrichment of FTO protein combined with photoaffinity labeling assay.
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| Enzyme Assay |
For non-cellular enzyme/receptor binding assays, the compound's inhibition of FTO demethylase activity can be assessed using biochemical assays. The FTO protein is incubated with a methylated RNA substrate and varying concentrations of 5-Aminofluorescein. The demethylation activity is measured, and the IC50 is calculated. The assay uses LC-MS analysis over a 1-hour incubation period. X-ray crystallography can be used to determine the binding mode.
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| Cell Assay |
For in vitro cell-based assays, cells are incubated with 5-Aminofluorescein-conjugated probes. Fluorescence intensity is measured using flow cytometry or fluorescence microscopy to assess labeling efficiency and target binding. The excitation wavelength is 490 nm and emission is 515 nm. For FTO inhibition studies, cells can be treated with 5-Aminofluorescein and m6A levels in mRNA are measured to assess demethylase inhibition.
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| Animal Protocol |
For in vivo imaging studies, 5-Aminofluorescein or its conjugates are administered to animal models. Fluorescence imaging is performed to track the distribution and accumulation of the labeled molecules. Excitation/emission at 490/515 nm is used for detection. For glioblastoma imaging, 5-Aminofluorescein conjugated to human serum albumin is used.
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| ADME/Pharmacokinetics |
Pharmacokinetic properties include solubility in DMF (5 mg/ml) and DMSO (5 mg/ml). The powder should be stored at -20°C, protected from light. It is stable at ambient temperature for a few days during shipping. The molecular weight is C20H13NO5. It has λmax of 226 nm.
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| Toxicity/Toxicokinetics |
As a fluorescent dye and prodrug component, its safety profile is generally considered acceptable for research applications. It is not intended for human use. No significant toxicity has been reported in standard research applications. The compound is a solid and should be handled with standard laboratory safety precautions.
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| References |
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| Additional Infomation |
5-Aminofluorescein is a primary amino compound, specifically fluorescein with an amino substituent at the C-5 position. It is a structural unit/intermediate in the synthesis of the fluorescent dye fluorescein; it can also be used to produce N-(fluorescein-5-yl)maleamic acid. Functionally, it is related to fluorescein (in its lactone form).
5-Aminofluorescein (5-AF) is a primary amino compound, specifically fluorescein with an amino substituent at the C-5 position. It is a structural unit/intermediate in the synthesis of the fluorescent dye fluorescein. It is functionally related to fluorescein (in its lactone form). Fluorescein derivatives can selectively inhibit FTO demethylation. The compound has been used for fluorescence imaging of glioblastomas and as a targeted prodrug. |
| Molecular Formula |
C20H13NO5
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|---|---|
| Molecular Weight |
347.33
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| Exact Mass |
347.079
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| CAS # |
3326-34-9
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| Related CAS # |
6-Aminofluorescein;51649-83-3
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| PubChem CID |
76845
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| Appearance |
Brown to reddish brown solid powder
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| Density |
1.6±0.1 g/cm3
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| Boiling Point |
695.3±55.0 °C at 760 mmHg
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| Melting Point |
223ºC
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| Flash Point |
374.3±31.5 °C
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| Vapour Pressure |
0.0±2.3 mmHg at 25°C
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| Index of Refraction |
1.820
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| LogP |
1.8
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
6
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| Rotatable Bond Count |
0
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| Heavy Atom Count |
26
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| Complexity |
557
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
GZAJOEGTZDUSKS-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C20H13NO5/c21-10-1-4-14-13(7-10)19(24)26-20(14)15-5-2-11(22)8-17(15)25-18-9-12(23)3-6-16(18)20/h1-9,22-23H,21H2
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| Chemical Name |
6-amino-3',6'-dihydroxyspiro[2-benzofuran-3,9'-xanthene]-1-one
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: This product requires protection from light (avoid light exposure) during transportation and storage. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ≥ 32 mg/mL (~92.13 mM)
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.8791 mL | 14.3955 mL | 28.7911 mL | |
| 5 mM | 0.5758 mL | 2.8791 mL | 5.7582 mL | |
| 10 mM | 0.2879 mL | 1.4396 mL | 2.8791 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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